TLR5 ligand induces the gene expression of antimicrobial peptides and CXCL8 through IL-1ß gene expression in cultured rumen epithelial cells.

High grain feeding or weaning, which could compromise the rumen epithelium by increasing ruminal short-chain fatty acid (SCFA) concentrations with pH reduction, is associated with high levels of ruminal toll-like receptor 5 (TLR5). This study aimed to determine the role of TLR5 in the rumen epithelium. Immunohistochemistry revealed that TLR5 was localized in cells on the basal side (i.e., basal and spinous layers) rather than in the granular layer in the rumen epithelium, where tight junctions are most potent, in pre- and post-weaning calves (n = 9). Primary bovine rumen epithelial cells (BRECs) obtained from Holstein cows (n = 3) were cultured to investigate the factors that upregulate TLR5; however, SCFA, low pH (pH 5.6), BHBA, L-lactate, D-lactate, and LPS did not upregulate TLR5 gene expression in BREC. Primary BREC treated with flagellin (TLR5 ligand) had higher expression of interleukin-1ß (IL-1ß) (P < 0.05) than BREC treated with vehicle. In addition, BREC treated with IL-1ß had higher expression of antimicrobial peptides and C-X-C motif chemokine ligand 8 than BREC treated with vehicle (P < 0.05). These results suggest that ruminal TLR5 may recognize epithelial disruption via flagellin and mediate the immune response via IL-1ß during high-grain feeding or weaning.

Effect of residual methane emission on physiological characteristics and carcass performance in Japanese Black cattle.

This study investigated the physiological characteristics and carcass performance associated with residual methane emissions (RME), and the effects of bull differences on CH4-related traits in Japanese Black cattle. Enteric methane (CH4) emissions from 156 Japanese Black cattle (111 heifers and 45 steers) were measured during early fattening using the sniffer method. Various physiological parameters were investigated to clarify the physiological traits between the high, middle, and low RME groups. CH4-related traits were examined to determine whether bull differences affected progeny CH4 emissions. Ruminal butyrate and NH3 concentrations were significantly higher in the high-RME group than in the low-RME group, whereas the propionate content was significantly higher in the low-RME group. Blood urea nitrogen, ß-hydroxybutyric acid, and insulin concentrations were significantly higher, and blood amino acids were lower in the high-RME group than in the other groups. No significant differences were observed in the carcass traits and beef fat composition between RME groups. CH4-related traits were significantly different among bull herds. Our results show that CH4-related traits are heritable, wherein bull differences affect progeny CH4 production capability, and that the above-mentioned rumen fermentations and blood metabolites could be used to evaluate enteric methanogenesis in Japanese Black cattle.

Advances in Methane Emission Estimation in Livestock: A Review of Data Collection Methods, Model Development and the Role of AI Technologies.

This review examines the significant role of methane emissions in the livestock industry, with a focus on cattle and their substantial impact on climate change. It highlights the importance of accurate measurement and management techniques for methane, a potent greenhouse gas accounting for 14-16% of global emissions. The study evaluates both conventional and AI-driven methods for detecting methane emissions from livestock, particularly emphasizing cattle contributions, and the need for region-specific formulas. Sections cover livestock methane emissions, the potential of AI technology, data collection issues, methane's significance in carbon credit schemes, and current research and innovation. The review emphasizes the critical role of accurate measurement and estimation methods for effective climate change mitigation and reducing methane emissions from livestock operations. Overall, it provides a comprehensive overview of methane emissions in the livestock industry by synthesizing existing research and literature, aiming to improve knowledge and methods for mitigating climate change. Livestock-generated methane, especially from cattle, is highlighted as a crucial factor in climate change, and the review underscores the importance of integrating precise measurement and estimation techniques for effective mitigation.

Assessing the impact of three feeding stages on rumen bacterial community and physiological characteristics of Japanese Black cattle.

In Japan, Japanese Black cattle, known for their exceptional meat quality owing to their abundant intramuscular fat, undergo a unique three-stage feeding system with varying concentrate ratios. There is limited research on physiological and rumen microbial changes in Japanese Black cattle during these stages. Therefore, this study aimed to examine Japanese Black steers in these three stages: early (T1, 12-14 months), middle (T2, 15-22 months), and late (T3, 23-30 months). The rumen bacteria of 21 cattle per phase was analyzed using 16S rRNA gene sequencing. Rumen bacterial diversity was significantly higher in T1, with a distinct distribution, than in T2 and T3. Specific phyla and genera were exclusive to each stage, reflecting the shifts in feed composition. Certain genera dominated each stage: T1 had Flexilinea, Streptococcus, Butyrivibrio, Selenomonas, and Kandleria; T2 had Bifidobacterium, Shuttleworthia, and Sharpea; and T3 had Acetitomaculum, Mycoplasma, Atopobium, and Howardella. Correlation analysis revealed significant associations between certain microbial populations and physiological parameters. These findings indicate that changes in energy content and feed composition are associated with physiological and ruminal alterations. This study may guide strategies to improve rumen health and productivity in Japanese Black cattle by modifying diets to specific fattening stages.

Effects of Rumen Fermentation Characteristics on Stress-Related Hormones and Behavior in Sheep.

This study investigated the relationship between rumen fermentation, stress-related hormones, and behavior in sheep with the aim of providing insights for improving animal welfare and feed management practices. Eight lambs were assigned to either a high concentration or control group. Blood samples were collected for hormone analysis, and an open field test was conducted to observe behavioral stress responses. The results showed that diet composition may affect the behavior of ruminants in response to stressors and novel situations, as evidenced by the higher number of escape attempts in the high-concentration group. In addition, analyses of individual volatile fatty acids (VFAs) showed a significant positive correlation between the acetic acid/propionic acid ratio and sniffing behavior of the novel object (p < 0.05, ρ = -0.414). These findings have important implications for animal welfare and feed management practices. Overall, this study provides insights into the potential impact of diet composition on the behavior of ruminants in response to stressors and novel situations, highlighting the importance of improving animal welfare through feed management practices. Further research is needed to fully elucidate the mechanisms underlying the complex relationship between rumen fermentation, stress-related hormones, and behavior in ruminant animals.

Physiological roles and regulation of hepatic angiopoietin-like protein 3 in Japanese Black cattle (Bos taurus) during the fattening period.

Angiopoietin-like protein 3 (ANGPTL3) is expressed predominantly in the liver and plays a major role in regulating the circulating triglyceride and lipoprotein fraction concentrations by inhibiting lipoprotein lipase (LPL) activity. Given these physiological roles, ANGPTL3 may play an important role in metabolic changes related to fat accumulation during the fattening period in Japanese Black. This study aimed to reveal the physiological roles of hepatic ANGPTL3 in Japanese Black steers (Bos taurus) during the fattening period and investigate the regulatory effects of hepatic ANGPTL3. To investigate the gene expression and protein localization of ANGPTL3, 18 tissue samples were collected from tree male Holstein bull calves aged 7 wk. Biopsied liver tissues and blood samples were collected from 21 Japanese Black steers during the early (T1; 13 mo of age), middle (T2; 20 mo), and late fattening phases (T3; 28 mo). Relative mRNA expression, blood metabolite concentrations, hormone concentrations, growth, and carcass traits were analyzed. To identify the regulatory factors of hepatic ANGPTL3, primary bovine hepatocytes collected by two Holstein calves aged 7 wk were incubated with insulin, palmitate, oleate, propionate, acetate, or beta-hydroxybutyric acid (BHBA). The ANGPTL3 gene was most highly expressed in the liver, with minor expression in the renal cortex, lungs, reticulum, and jejunum in Holstein bull calves. In Japanese Black steers, relative ANGPTL3 mRNA expressions were less as fattening progressed, and blood triglyceride, total cholesterol, and nonesterified fatty acid (NEFA) concentrations increased. Relative ANGPTL8 and Liver X receptor alpha (LXRα) mRNA expressions decreased in late and middle fattening phases, respectively. Furthermore, relative ANGTPL3 mRNA expression was positively correlated with ANGPTL8 (r = 0.650; P < 0.01) and ANGPTL4 (r = 0.540; P < 0.05) in T3 and T1, respectively, and LXRα showed no correlation with ANGPTL3. Relative ANGTPL3 mRNA expression was negatively correlated with total cholesterol (r = -0.434; P < 0.05) and triglyceride (r = -0.645; P < 0.01) concentrations in T3 and T1, respectively; There was no significant correlation between ANGTPL3 and carcass traits. Relative ANGTPL3 mRNA expression in cultured bovine hepatocytes was downregulated in oleate treatment. Together, these findings suggest that ANGPTL3 downregulation in late fattening phases is associated with the changes in lipid metabolism.

The role of angiopoietin-like protein 3 (ANGPTL3) in various animal species under different physiological conditions remains largely unknown. We evaluated the physiological roles of hepatic ANGPTL3 in Japanese Black steers (Bos taurus) during the fattening period and investigated the expressional regulation of ANGPTL3 in bovine hepatocytes. Relative ANGPTL3 mRNA expression decreased late in the fattening phases. Relative ANGPTL3 mRNA expression was positively correlated with ANGPTL4 and ANGPTL8 and was negatively correlated with blood triglyceride concentrations in early fattening phases. Relative ANGPTL3 mRNA expression in cultured bovine hepatocytes was downregulated in oleate treatment. Fatty acids may influence ANGPTL3 expression in cultured bovine hepatocytes through possible regulatory factors. Our findings suggest that the physiological roles of ANGPTL3 are associated with the changes of lipid metabolism during the fattening period, and the ANGPTL family seem to be associated with blood lipid metabolites.

Characteristics of Physiological Parameters of Japanese Black Calves Relate to Carcass Weight.

This study aimed to identify the growth performance and blood factors associated with carcass weight in Japanese Black calves based on 675 performance tests and field carcass records. We measured the body weight, withers height, and chest girth at the start of fattening age (approximately 8-10 months) and analyzed eight blood factors, including vitamins and metabolites. Single- and two-trait animal models were used to estimate the heritability and genetic correlations. The heritability estimates for growth performance were moderate to high (ranging from 0.48 to 0.74), and those for blood metabolites were low to moderate (ranging from 0.19 to 0.51). Estimates for genetic correlations of carcass or body weight with body weight, withers height, and chest girth were high (ranging from 0.42 to 0.80). The body weight and withers height at 8 months of age are possibly closely related to the final carcass weight. The blood metabolites associated with body weight were vitamin E in steers (castrated males) and ß-carotene in heifers. Our findings indicate that body measurements and blood metabolites measured during the growing period could be used to determine the nutritional and physiological status of cattle as well as predict carcass weight.

Expression of chemerin in intestinal mucosa of calves with comparable expression level with other antimicrobial proteins.

Neonatal calves are highly susceptible to infectious disorders including diarrhea. Therefore, epithelial innate immunity, including antimicrobial peptides/proteins (AMPs), is important during the early stage of their lives. Chemerin, a multifunctional protein that was originally identified as a chemokine, possesses a potent antimicrobial activity. The present study investigated the expression levels of chemerin in the gastrointestinal (GI) tract of growing calves. Chemerin and its coding gene, retinoic acid receptor responder protein 2 (RARRES2), were highly expressed in duodenum, jejunum, and ileum compared with other parts of the GI tract. Immunohistochemistry demonstrated that chemerin-producing cells were localized in the crypt of the intestinal mucosa. Finally, the expression level of RARRES2 was higher compared with those of other major AMPs in duodenum, although it was lower compared with that of enteric ß-defensin but mostly higher than those of other AMPs in jejunum and ileum at various ages in calves. The expression levels of RARRES2 were not influenced by the age of calves in duodenum and jejunum, whereas a higher expression level of RARRES2 in ileum was observed in younger calves. This study revealed that chemerin is produced in the small intestine of calves and has the potential to contribute to the gut epithelial barrier system.

Physiological responses and adaptations to high methane production in Japanese Black cattle.

In this study, using enteric methane emissions, we investigated the metabolic characteristics of Japanese Black cattle. Their methane emissions were measured at early (age 13 months), middle (20 months), and late fattening phases (28 months). Cattle with the highest and lowest methane emissions were selected based on the residual methane emission values, and their liver transcriptome, blood metabolites, hormones, and rumen fermentation characteristics were analyzed. Blood ß-hydroxybutyric acid and insulin levels were high, whereas blood amino acid levels were low in cattle with high methane emissions. Further, propionate and butyrate levels differed depending on the enteric methane emissions. Hepatic genes, such as SERPINI2, SLC7A5, ATP6, and RRAD, which were related to amino acid transport and glucose metabolism, were upregulated or downregulated during the late fattening phase. The above mentioned metabolites and liver transcriptomes could be used to evaluate enteric methanogenesis in Japanese Black cattle.

Changes in the liver transcriptome and physiological parameters of Japanese Black steers during the fattening period.

We investigated the physiological changes during the fattening period and production characteristics in Japanese Black steers bred and raised using the typical feeding system in Japan. Here, 21 Japanese Black steers aged 12 months were used, with experimental period divided into early (12-14 months of age), middle (15-22 months), and late fattening phases (23-30 months). The liver transcriptome, blood metabolites, hormones, and rumen fermentation characteristics were analyzed. Blood triglyceride and non-esterified fatty acid concentrations increased, whereas blood ketone levels decreased, with fattening phases. Blood insulin increased with fattening phases and was positively correlated with carcass weight and marbling in late fattening phases. Rumen fermentation characteristics showed high propionate levels and low butyrate levels in late fattening phases, likely due to increased energy intake. Genes related to glucose metabolism, such as SESN3, INSR, LEPR, and FOXO3, were down-regulated in late fattening phases. Genes related to lipid metabolism, such as FABP4, were up-regulated, whereas FADS1 and FADS2 were down-regulated. These findings suggest that the physiological changes resulted from changes in the energy content and composition of diets. Liver metabolism changed with changes in fat metabolism. Insulin was strongly associated with physiological changes and productivity in Japanese Black cattle.

Maternal Nutrient Restriction Disrupts Gene Expression and Metabolites Associated with Urea Cycle, Steroid Synthesis, Glucose Homeostasis, and Glucuronidation in Fetal Calf Liver.

This study aimed to understand the mechanisms underlying the effects of maternal undernutrition (MUN) on liver growth and metabolism in Japanese Black fetal calves (8.5 months in utero) using an approach that integrates metabolomics and transcriptomics. Dams were fed 60% (low-nutrition; LN) or 120% (high-nutrition; HN) of their overall nutritional requirements during gestation. We found that MUN markedly decreased the body and liver weights of the fetuses; metabolomic analysis revealed that aspartate, glycerol, alanine, gluconate 6-phosphate, and ophthalmate levels were decreased, whereas UDP-glucose, UDP-glucuronate, octanoate, and 2-hydroxybutyrate levels were decreased in the LN fetal liver (p ≤ 0.05). According to metabolite set enrichment analysis, the highly different metabolites were associated with metabolisms including the arginine and proline metabolism, nucleotide and sugar metabolism, propanoate metabolism, glutamate metabolism, porphyrin metabolism, and urea cycle. Transcriptomic and qPCR analyses revealed that MUN upregulated QRFPR and downregulated genes associated with the glucose homeostasis (G6PC, PCK1, DPP4), ketogenesis (HMGCS2), glucuronidation (UGT1A1, UGT1A6, UGT2A1), lipid metabolism (ANGPTL4, APOA5, FADS2), cholesterol and steroid homeostasis (FDPS, HSD11B1, HSD17B6), and urea cycle (CPS1, ASS1, ASL, ARG2). These metabolic pathways were extracted as relevant terms in subsequent gene ontology/pathway analyses. Collectively, these results indicate that the citrate cycle was maintained at the expense of activities of the energy metabolism, glucuronidation, steroid hormone homeostasis, and urea cycle in the liver of MUN fetuses.

Genetic relationships of feed efficiency and growth traits with carcass traits in Japanese Shorthorn cattle.

In this study, we examined genetic parameters for feed efficiency, growth, and carcass traits in Japanese Shorthorn cattle, based on 714 performance tests and 15,790 field carcass records. Feed efficiency traits, including residual feed intake (RFI) and residual body weight gain (RG), were calculated. Single-trait and two-trait animal models were used to estimate heritability and genetic correlations. Heritability estimates for feed efficiency traits were found to be low to moderate (ranging from 0.03 to 0.36); notably, heritability was moderate for RG and low for RFI. Estimates for genetic correlations between feed efficiency traits and average daily gain (DG) were favorably moderate to high (absolute values of 0.43-0.85), and those with daily feed intake were low (absolute values of 0.00-0.32). We also estimated a high genetic correlation between RG and DG. The backfat thickness (BF) of bull calves showed favorable or no genetic correlation estimates with feed efficiency and growth traits, whereas RG and BF showed favorable or no genetic correlation estimates with carcass traits. Our findings indicate that genetic improvements in both feed utilization ability and carcass traits could be achieved by utilizing RG and BF in Japanese Shorthorn cattle.

Chemerin Regulates Epithelial Barrier Function of Mammary Glands in Dairy Cows.

Epithelial barrier function in the mammary gland acts as a forefront of the defense mechanism against mastitis, which is widespread and a major disorder in dairy production. Chemerin is a chemoattractant protein with potent antimicrobial ability, but its role in the mammary gland remains unelucidated. The aim of this study was to determine the function of chemerin in mammary epithelial tissue of dairy cows in lactation or dry-off periods. Mammary epithelial cells produced chemerin protein, and secreted chemerin was detected in milk samples. Chemerin treatment promoted the proliferation of cultured bovine mammary epithelial cells and protected the integrity of the epithelial cell layer from hydrogen peroxide (H2O2)-induced damage. Meanwhile, chemerin levels were higher in mammary tissue with mastitis. Tumor necrosis factor alpha (TNF-α) strongly upregulated the expression of the chemerin-coding gene (RARRES2) in mammary epithelial cells. Therefore, chemerin was suggested to support mammary epithelial cell growth and epithelial barrier function and to be regulated by inflammatory stimuli. Our results may indicate chemerin as a novel therapeutic target for diseases in the bovine mammary gland.

Evaluation of inbreeding and genetic diversity in Japanese Shorthorn cattle by pedigree analysis.

The Japanese Shorthorn is a Japanese Wagyu breed maintained at a small population size. We assessed the degree of inbreeding and genetic diversity among Japanese Shorthorn cattle using pedigree analysis. We analyzed the pedigree records of registered Japanese Shorthorn born between 1980 and 2018, after evaluating the pedigree completeness. The average of the actual inbreeding coefficients increased at the same rates annually from approximately 1.5% in 1980 to 4.2% in 2018 and was higher than the expected inbreeding coefficients over time. The effective population size based on the individual coancestry rate largely decreased from 127.8 in 1980 to 82.6 in 1999, and then remained almost constant at approximately 90. Three effective numbers of ancestors decreased over time until 1995, then remained almost constant. In particular, the effective number of founder genomes (Nge ) decreased from 43.8 in 1980 to 11.9 in 2018. The index of genetic diversity based on Nge decreased from 0.99 in 1980 to 0.96 in 2018 due to genetic drift in non-founder generations. Changes in inbreeding and genetic diversity parameters were similar between Japanese Shorthorn and other Japanese Wagyu breeds, but the magnitude of the changes was lower in the Japanese Shorthorn.

Maternal Undernutrition during Pregnancy Alters Amino Acid Metabolism and Gene Expression Associated with Energy Metabolism and Angiogenesis in Fetal Calf Muscle.

To elucidate the mechanisms underlying maternal undernutrition (MUN)-induced fetal skeletal muscle growth impairment in cattle, the longissimus thoracis muscle of Japanese Black fetal calves at 8.5 months in utero was analyzed by an integrative approach with metabolomics and transcriptomics. The pregnant cows were fed on 60% (low-nutrition, LN) or 120% (high-nutrition, HN) of their overall nutritional requirement during gestation. MUN markedly decreased the bodyweight and muscle weight of the fetus. The levels of amino acids (AAs) and arginine-related metabolites including glutamine, gamma-aminobutyric acid (GABA), and putrescine were higher in the LN group than those in the HN group. Metabolite set enrichment analysis revealed that the highly different metabolites were associated with the metabolic pathways of pyrimidine, glutathione, and AAs such as arginine and glutamate, suggesting that MUN resulted in AA accumulation rather than protein accumulation. The mRNA expression levels of energy metabolism-associated genes, such as PRKAA1, ANGPTL4, APLNR, CPT1B, NOS2, NOS3, UCP2, and glycolytic genes were lower in the LN group than in the HN group. The gene ontology/pathway analysis revealed that the downregulated genes in the LN group were associated with glucose metabolism, angiogenesis, HIF-1 signaling, PI3K-Akt signaling, pentose phosphate, and insulin signaling pathways. Thus, MUN altered the levels of AAs and expression of genes associated with energy expenditure, glucose homeostasis, and angiogenesis in the fetal muscle.

Effects of low and high levels of maternal nutrition consumed for the entirety of gestation on the development of muscle, adipose tissue, bone, and the organs of Wagyu cattle fetuses.

This study aimed to investigate the effects of high and low levels of energy intake during the entire gestation period on the skeletal muscle development, organ development, and adipose tissue accumulation in fetuses of Wagyu (Japanese Black) cows, a breed with highly marbled beef. Cows were allocated to a high-nutrition (n = 6) group (fed 120% of the nutritional requirement) or low-nutrition (n = 6) group (fed 60% of the nutritional requirement). The cows were artificially inseminated with semen from the same sire, and the fetuses were removed by cesarean section at 260 ± 8.3 days of fetal age and slaughtered. The whole-body, total muscle, adipose, and bone masses of the fetal half-carcasses were significantly higher in the high-nutrition group than the low-nutrition group (p = 0.0018, 0.009, 0.0004, and 0.0362, respectively). Fifteen of 20 individual muscles, five of six fat depots, nine of 17 organs, and seven of 12 bones that were investigated had significantly higher masses in the high-nutrition group than the low-nutrition group. The crude components and amino acid composition of the longissimus muscle significantly differed between the low- and high-nutrition groups. These data indicate that maternal nutrition during gestation has a marked effect on the muscle, bone, and adipose tissue development of Wagyu cattle fetuses.

The Physiological Roles of Vitamin E and Hypovitaminosis E in the Transition Period of High-Yielding Dairy Cows.

Levels of alpha-tocopherol (α-Toc) decline gradually in blood throughout prepartum, reaching lowest levels (hypovitaminosis E) around calving. Despite numerous reports about the disease risk in hypovitaminosis E and the effect of α-Toc supplementation on the health of transition dairy cows, its risk and supplemental effects are controversial. Here, we present some novel data about the disease risk of hypovitaminosis E and the effects of α-Toc supplementation in transition dairy cows. These data strongly demonstrate that hypovitaminosis E is a risk factor for the occurrence of peripartum disease. Furthermore, a study on the effectiveness of using serum vitamin levels as biomarkers to predict disease in dairy cows was reported, and a rapid field test for measuring vitamin levels was developed. By contrast, evidence for how hypovitaminosis E occurred during the transition period was scarce until the 2010s. Pioneering studies conducted with humans and rodents have identified and characterised some α-Toc-related proteins, molecular players involved in α-Toc regulation followed by a study in ruminants from the 2010s. Based on recent literature, the six physiological factors: (1) the decline in α-Toc intake from the close-up period; (2) changes in the digestive and absorptive functions of α-Toc; (3) the decline in plasma high-density lipoprotein as an α-Toc carrier; (4) increasing oxidative stress and consumption of α-Toc; (5) decreasing hepatic α-Toc transfer to circulation; and (6) increasing mammary α-Toc transfer from blood to colostrum, may be involved in α-Toc deficiency during the transition period. However, the mechanisms and pathways are poorly understood, and further studies are needed to understand the physiological role of α-Toc-related molecules in cattle. Understanding the molecular mechanisms underlying hypovitaminosis E will contribute to the prevention of peripartum disease and high performance in dairy cows.

Forestomach developmental failure in an 11-month-old Japanese Black steer with severely retarded growth and chronic ruminal tympany.

This study reports findings from the pathological examination of the forestomach of an 11-month-old Japanese Black steer with severely retarded growth (41% of expected weight) and chronic ruminal tympany. The ruminal papillae were weakly formed (0.3-0.5 cm long) and unevenly distributed. The cellulae and cristae reticuli were underdeveloped; the cristae were 0.4-0.7 cm in height and milky white. The keratinized layer in the stratified squamous epithelium was thickened. Ruminal pH was 5.25, and ruminal volatile fatty acid concentration was 11.7 mM. The steer's severely retarded growth was considered to be caused by malnutrition due to developmental and functional failure of the forestomach.

Maternal Nutrition During Gestation Alters Histochemical Properties, and mRNA and microRNA Expression in Adipose Tissue of Wagyu Fetuses.

We hypothesized that maternal low or high nutrition would give unique effects to morphological and molecular dynamics in adipose tissue of fetus of fatty breed Wagyu (Japanese Black) cattle which produce highly marbled beef. This study aimed to determine the effects of maternal energy intake in Wagyu cows, during gestation on fetal adipose tissue development, histochemical properties, and gene and microRNA (miRNA) expression. Cows were allocated to one of two nutritional energy groups: 120% (HIGH) or 60% nutritional requirements of (LOW). Fetuses (n = 6 per treatment) were removed from pregnant cows by cesarean section at fetal age 260 ± 8 days and euthanized. Subcutaneous adipose tissue (SAT), thoracic cavity visceral adipose tissue (TVAT), and perirenal adipose tissue (PAT) were collected for analysis. In histochemical analysis, in SAT and PAT, HIGH fetuses had greater diameter of adipocytes than LOW fetuses (P<0.05). Only in SAT, LOW fetuses had more Leptin (LEP) mRNA and tended to have more Peroxisome Proliferator-Activated Receptor gamma (PPARG) CCAAT-enhancer-binding proteins alpha (CEBPA) and Glucose transporter (GLUT) 4 mRNA(P<0.10). In all SAT, TVAT, and PAT, LOW fetuses had higher levels of the brown adipose tissue (BAT) biomarkers Uncoupling Protein (UCP) 1 and PPARG coactivator (PGC) 1α mRNA than HIGH fetuses (P<0.08). Meanwhile, in the other adipose tissue, LOW fetuses had lower PPARG, CEBPA, and Zinc Finger Protein (ZFP) 423 (in TVAT and PAT), FASN (in TVAT), LEP and GLUT4 mRNA (in PAT; P<0.10). In particular, in TVAT and PAT, LOW fetuses exhibited lower expression of WAT biomarkers (PPARG and ZFP423). Differential expression of various miRNAs related to adipogenesis between the LOW and HIGH fetuses was detected in an adipose tissue-specific manner (P<0.10). Based on adipose tissue-specific effects of maternal nutrition, these findings suggested that poor maternal nutrition in Wagyu cattle increased BAT development in SAT, TVAT and PAT, while elevated maternal nutrition stimulated fetal SAT development compared with that of TVAT and PAT.

Development of multipurpose recombinant reporter bovine leukemia virus.

Bovine leukemia virus (BLV) is a global problem that results in significant economic losses to the livestock industry. We developed three virus strains by inserting the HiBiT reporter tag from NanoLuc luciferase (NLuc) into limited sites within BLV molecular clones. Initial analysis for site selection of the tag insertion revealed a permissible site immediately downstream of the viral envelope gene. Therefore, NLuc activity could be used to measure virus copy numbers in the supernatant and the levels of cell infection. Productivity and growth kinetics of the reporter virus were similar to those of the wild-type strain; therefore, the reporter virus can be used to characterize the replication of chimeric viruses as well as responses to the antiviral drug, amprenavir. Collectively, our results suggest that the BLV reporter virus with a HiBiT tag insertion is a highly versatile system for various purposes such as evaluating virus replication and antiviral drugs.